We intend to study the protein-nucleic acid interactions which determine the specificity of initiation of the transcription and translation of messenger RNAs in bacteria. Previous work has provided 1) some knowledge of the oligonucleotide sequences which are recognized when RNA polymerases begin synthesis or when ribosomes initiate a polypeptide chain; and 2) the realization that alteration of these recognition phenomena is important to both normal cellular development and the success of viral infection. However we still lack a detailed understanding of how ribosomes and polymerases recognize start signals and how control over gene expression is exerted at these points. The molecular mechanism of transcriptional and translational initiation will be examined using three bacteriophage systems. 1) The RNA bacteriophage messengers, because they are well-characterized and are available in large amounts, will be utilized for physical studies of RNA secondary/tertiary structure and for further analysis of specificity of ribosomes, initiation and interference factors, and other RNA-binding proteins. 2) Bacteriophage T7 provides two discrete sets of stable, intracellular mRNAs, which can be studied after transcription by their respective polymerases (E. coli and T7) either in vivo or in vitro; their synthesis and translation appear to be controlled by both viral and host functions (which differ in male and female cells). 3) The lactose repressor mRNA will be synthesized in vitro from lac transducing phage DNA; it has been the subject of intensive genetic manipulation, which provides not only several promoter mutations, but also translational restarts at new locations in the RNA. In each case, nucleotide sequences of the RNA termini, the protein synthesis initiator regions, and sites bound by other proteins will be determined by Sanger methods. Alterations in recognition after viral infection will be correlated with changes in cellular components. Particular emphasis will be placed on elucidating the role of mRNA secondary and tertiary structure in translational control. BIBLIOGRAPHIC REFERENCES: M. Rosenberg, R.A. Kramer and J.A. Steitz, "The Specificity of RNase III Cleavage of Bacteriophage T7 Early Messenger RNAs", Brookhaven Symposium (1975) Vol. 26, 277-285.